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Intrinsic and extrinsic factors, including lifestyle and sun exposure, can contribute to cell senescence, which impairs skin homeostasis, that may in turn lead to skin aging. Senescent cells have a specific secretome, called the senescence-associated secretory phenotype (SASP) that includes MMPs, CXCLs and S100A8/9. Reducing the SASP with senotherapeutics is a promising strategy to reduce skin aging. Here we evaluated the effect of a formula containing niacinamide and hyaluronic acid, which are known to limit senescence and skin aging. We conducted three different studies. (1) Ex vivo explants treated with the formula had more collagen and glycosaminoglycan. (2) In a clinical trial with forty-four women, two months of treatment improved fine lines, wrinkles, luminosity, smoothness, homogeneity, and plumpness. (3) In a third study on thirty women, we treated one arm for two months and took skin biopsies to study gene expression. 101 mRNAs and 13 miRNAs were differentially expressed. We observed a likely senomorphic effect, as there was a decrease in many SASP genes including MMP12 and CXCL9 and a significant downregulation of autocrine signaling genes: S100A8 and S100A9. These pharmaco-clinical results are the first to demonstrate the senomorphic properties of an effective anti-aging formula in skin.
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Ácido Hialurónico , Niacinamida , Envejecimiento de la Piel , Humanos , Ácido Hialurónico/farmacología , Ácido Hialurónico/metabolismo , Envejecimiento de la Piel/efectos de los fármacos , Femenino , Persona de Mediana Edad , Niacinamida/farmacología , Adulto , Fenotipo Secretor Asociado a la Senescencia , Piel/efectos de los fármacos , Piel/metabolismo , Piel/patología , Senescencia Celular/efectos de los fármacos , AncianoRESUMEN
RAS proteins (KRAS, NRAS and HRAS) are frequently activated in different cancer types (e.g., non-small cell lung cancer, colorectal cancer, melanoma and bladder cancer). For many years, their activities were considered redundant due to their high degree of sequence homology (80% identity) and their shared upstream and downstream protein partners. However, the high conservation of the Hyper-Variable-Region across mammalian species, the preferential activation of different RAS proteins in specific tumor types and the specific post-translational modifications and plasma membrane-localization of each paralog suggest they could ensure discrete functions. To gain insights into RAS proteins specificities, we explored their proximal protein-protein interaction landscapes using the proximity-dependent biotin identification technology (BioID) in Flp-In T-REx 293 cell lines stably transfected and inducibly expressing wild type KRAS4B, NRAS or HRAS. We identified more than 800 high-confidence proximal interactors, allowing us to propose an unprecedented comparative analysis of wild type RAS paralogs protein networks. These data bring novel information on poorly characterized RAS functions, e.g., its putative involvement in metabolic pathways, and on shared as well as paralog-specific protein networks that could partially explain the complexity of RAS functions. These networks of protein interactions open numerous avenues to better understand RAS paralogs biological activities.
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Although the development of mitogen-activated protein kinase (MAPK) inhibitors has greatly improved the prognosis of BRAFV600 cutaneous melanomas, the identification of molecular indicators for mutated patients at risk of early progression remains a major issue. Using an amplicon-based next-generation-sequencing (NGS) assay that targets cancer-related genes, we investigated co-occurring alterations in 89 melanoma samples. We analyzed both their association with clinicopathological variables and clinical significance in terms of progression-free survival (PFS) and overall survival (OS) according to BRAF genotyping. Among co-occurring mutations, TERT promoter was the most frequently mutated gene. Although no significant difference in PFS was observed in the presence or absence of co-occurring alterations to BRAFV600, there was a trend of longer PFS for patients harboring TERT c.-124C>T mutation. Of most interest, this mutation is an independent marker of good prognosis in subgroups of patients with poor prognosis (presence of brain metastasis and elevated level of lactate dehydrogenase, LDH). Moreover, combination of elevated LDH level, presence of brain metastasis, and TERT c.-124C>T mutation was identified as the best fit model for predicting clinical outcome. Our work revealed the potential interest of c.-124C>T status determination in order to refine the prognosis of BRAFV600 melanoma under mitogen-activated protein kinase (MAPK) inhibitors.
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Purpose: Among the FKBP family members, FKBP4 has been described to have a potential role in tumorigenesis, and as a putative tissue marker. We previously showed that FKBP4, an HSP90-associated co-chaperone, can elicit immune response as a tumor-specific antigen, and are overexpressed in breast cancer. Experimental design: In this study, we examined how loss of FKBP4 affect breast cancer progression and exploited protein interactomics to gain mechanistic insight into this process. Results: We found that FKBP4 expression is associated with breast cancer progression and prognosis, especially of ER-negative breast cancer. Furthermore, FKBP4 depletion specifically reduces cell growth and proliferation of triple negative breast cancer cell model and xenograft tumor model. Using specific protein interactome strategy by BirA proximity-dependent biotin identification, we demonstrated that FKBP4 is a novel PI3K-Akt-mTOR proximal interacting protein. Conclusion: Our results suggest that FKBP4 interacts with PI3K and can enhance Akt activation through PDK1 and mTORC2.
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Proteínas Quinasas Dependientes de 3-Fosfoinosítido/metabolismo , Neoplasias de la Mama/metabolismo , Diana Mecanicista del Complejo 2 de la Rapamicina/metabolismo , Fosfatidilinositol 3-Quinasas/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Proteínas de Unión a Tacrolimus/metabolismo , Proteínas Quinasas Dependientes de 3-Fosfoinosítido/genética , Animales , Neoplasias de la Mama/genética , Línea Celular Tumoral , Proliferación Celular , Femenino , Humanos , Diana Mecanicista del Complejo 2 de la Rapamicina/genética , Ratones Desnudos , Fosfatidilinositol 3-Quinasas/genética , Unión Proteica , Proteínas Proto-Oncogénicas c-akt/genética , Transducción de Señal , Proteínas de Unión a Tacrolimus/genéticaRESUMEN
Introduction: The proteome is a dynamic system in which protein-protein interactions play a crucial part in shaping the cell phenotype. However, given the current limitations of available technologies to describe the dynamic nature of these interactions, the identification of protein-protein interactions has long been a major challenge in proteomics. In recent years, the development of BioID and APEX, two proximity-tagging technologies, have opened-up new perspectives and have already started to change our conception of protein-protein interactions, and more generally, of the proteome. With a broad range of application encompassing health, these new technologies are currently setting milestones crucial to understand fine cellular mechanisms. Area covered: In this article, we describe both the recent and the more conventional available tools to study protein-protein interactions, compare the advantages and the limitations of these techniques, and discuss the recent advancements led by the proximity tagging techniques to refine our conception of the proteome. Expert opinion: The recent development of proximity labeling techniques emphasizes the growing importance of such technologies to decipher cellular mechanism. Although several challenges still need to be addressed, many fields can benefit from these tools and notably the detection of new therapeutic targets for patient care.
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Proteómica/métodos , Coloración y Etiquetado/métodos , Animales , Humanos , Espectrometría de Masas , Unión ProteicaRESUMEN
The proteome is a dynamic system in which protein-protein interactions play a crucial role to model together the cellular phenotype. However, given the inherent limitation of the available technologies to depict the dynamic nature of these interactions, identify protein-protein interaction has for a long time represented an important challenge in proteomic. The recent development of BioID and APEX, two proximity-dependent labeling technologies, opens today new perspectives and yet start changing our vision of protein-protein interaction, and more globally our vision of the proteome. In this review, we describe the recent and conventional tools available to study protein-protein interactions, compare the advantages and limitations of these technics, and discuss the recent progress brought by the proximity-dependent labelling to complete our vision of the proteome, and thus better understand cellular mechanisms.
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Mapas de Interacción de Proteínas , Proteómica/métodos , Coloración y Etiquetado/métodos , Animales , Biotinilación/métodos , Humanos , Unión Proteica , Dominios y Motivos de Interacción de Proteínas , Proteoma/análisisRESUMEN
Rearrangements of the anaplastic lymphoma kinase (ALK) gene in non-small cell lung cancer (NSCLC) represent a novel molecular target in a small subset of tumors. Although ALK rearrangements are usually assessed by immunohistochemistry (IHC) and fluorescence in situ hybridization (FISH), molecular approaches have recently emerged as relevant alternatives in routine laboratories. Here, we evaluated the use of two different amplicon-based next-generation sequencing (NGS) methods (AmpliSeq and Archer®FusionPlex®) to detect ALK rearrangements, and compared these with IHC and FISH. A total of 1128 NSCLC specimens were screened using conventional analyses, and a subset of 37 (15 ALK-positive, and 22 ALK-negative) samples were selected for NGS assays. Although AmpliSeq correctly detected 25/37 (67.6%) samples, 1/37 (2.7%) and 11/37 (29.7%) specimens were discordant and uncertain, respectively, requiring further validation. In contrast, Archer®FusionPlex® accurately classified all samples and allowed the correct identification of one rare DCTN1-ALK fusion, one novel CLIP1-ALK fusion, and one novel GCC2-ALK transcript. Of particular interest, two out of three patients harboring these singular rearrangements were treated with and sensitive to crizotinib. These data show that Archer®FusionPlex® may provide an effective and accurate alternative to FISH testing for the detection of known and novel ALK rearrangements in clinical diagnostic settings.
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Adenocarcinoma del Pulmón/genética , Quinasa de Linfoma Anaplásico/genética , Carcinoma de Pulmón de Células no Pequeñas/genética , Secuenciación de Nucleótidos de Alto Rendimiento/métodos , Neoplasias Pulmonares/genética , Proteínas de Fusión Oncogénica/genética , Adenocarcinoma del Pulmón/tratamiento farmacológico , Adenocarcinoma del Pulmón/metabolismo , Adenocarcinoma del Pulmón/cirugía , Anciano , Quinasa de Linfoma Anaplásico/metabolismo , Antineoplásicos/uso terapéutico , Carcinoma de Pulmón de Células no Pequeñas/tratamiento farmacológico , Carcinoma de Pulmón de Células no Pequeñas/metabolismo , Carcinoma de Pulmón de Células no Pequeñas/cirugía , Estudios de Casos y Controles , Crizotinib/uso terapéutico , Complejo Dinactina/genética , Complejo Dinactina/metabolismo , Femenino , Expresión Génica , Proteínas de la Matriz de Golgi/genética , Proteínas de la Matriz de Golgi/metabolismo , Secuenciación de Nucleótidos de Alto Rendimiento/instrumentación , Humanos , Hibridación Fluorescente in Situ , Neoplasias Pulmonares/tratamiento farmacológico , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/cirugía , Masculino , Proteínas Asociadas a Microtúbulos/genética , Proteínas Asociadas a Microtúbulos/metabolismo , Persona de Mediana Edad , Estadificación de Neoplasias , Proteínas de Fusión Oncogénica/metabolismo , ARN Mensajero/genética , ARN Mensajero/metabolismoRESUMEN
Pore-forming toxins are potent virulence factors secreted by a large array of bacteria. Here, we deciphered the action of ExlA from Pseudomonas aeruginosa and ShlA from Serratia marcescens on host cell-cell junctions. ExlA and ShlA are two members of a unique family of pore-forming toxins secreted by a two-component secretion system. Bacteria secreting either toxin induced an ExlA- or ShlA-dependent rapid cleavage of E-cadherin and VE-cadherin in epithelial and endothelial cells, respectively. Cadherin proteolysis was executed by ADAM10, a host cell transmembrane metalloprotease. ADAM10 activation is controlled in the host cell by cytosolic Ca2+ concentration. We show that Ca2+ influx, induced by ExlA or ShlA pore formation in the plasma membrane, triggered ADAM10 activation, thereby leading to cadherin cleavage. Our data suggest that ADAM10 is not a cellular receptor for ExlA and ShlA, further confirming that ADAM10 activation occurred via Ca2+ signalling. In conclusion, ExlA- and ShlA-secreting bacteria subvert a regulation mechanism of ADAM10 to activate cadherin shedding, inducing intercellular junction rupture, cell rounding and loss of tissue barrier integrity.
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Proteína ADAM10/metabolismo , Proteínas Bacterianas/metabolismo , Cadherinas/metabolismo , Infecciones por Bacterias Gramnegativas/metabolismo , Proteínas Hemolisinas/metabolismo , Animales , Toxinas Bacterianas/metabolismo , Western Blotting , Calcio/metabolismo , Activación Enzimática , Femenino , Humanos , Ratones , Ratones Endogámicos BALB C , Microscopía Confocal , Pseudomonas aeruginosa/patogenicidad , Serratia marcescens/patogenicidad , Virulencia/fisiología , Factores de Virulencia/metabolismoRESUMEN
Circulating tumoral DNA (ctDNA), commonly named "liquid biopsy", has emerged as a new promising noninvasive tool to detect biomarker in several cancers including lung cancer. Applications involving molecular analysis of ctDNA in lung cancer have increased and encompass diagnosis, response to treatment, acquired resistance and prognosis prediction, while bypassing the problem of tumor heterogeneity. ctDNA may then help perform dynamic genetic surveillance in the era of precision medicine through indirect tumoral genomic information determination. The aims of this review were to examine the recent technical developments that allowed the detection of genetic alterations of ctDNA in lung cancer. Furthermore, we explored clinical applications in patients with lung cancer including treatment efficiency monitoring, acquired therapy resistance mechanisms and prognosis value.